Glaucoma is a prevalent blinding disease characterized by the progressive loss of retinal ganglion cells. Previously, we used Bax knockout mice to show that this proapoptotic gene was essential for ganglion cell death stimulated by optic nerve crush and in a mouse model of spontaneous glaucoma. During this work, we found that mice heterozygous for the Bax knock out allele have dramatically different phenotypes depending on their genetic background. In some lines, Bax+/- mice have a wild type phenotype, (all ganglion cells die), while in other lines, Bax+/- mice have a mutant phenotype, (all the cells survive). This proposal will extend our studies on the effects of Bax in retinal ganglion cells in an optic nerve crush model. In the first aim, we will examine the basis for the variable effects of the Bax+/- genotype in different lines. Preliminary studies indicate that two distinct lines (DBA/2J and 129B6, which have all-or-none phenotypes as Bax heterozygotes), exhibit different levels of Bax gene expression (high and low levels), consistent with their phenotype. This phenomenon will be studied further by analyzing the sequence and strength of the Bax promoters in each line. Axon loss precedes soma death in glaucoma. In the second Aim, we will assess if regeneration can be stimulated in cells arrested in the cell death pathway at the level of Bax. Regeneration will be stimulated in Bax+/+ and Bax-/- ganglion cells using established methods to determine if the Bax deficient cells have a greater capacity for new growth. These experiments are critical to show that blocking cell death is practical, since a live cell with no axon is still non-functional. To complete the strategy of targeting Bax, in the third Aim we will examine several reagents that are designed to reduce Bax expression and/or function in wild type ganglion cells. These reagents interfere with either BAX translation, the cellular translocation of BAX to mitochondria, or prevent the down stream effects of BAX. Ultimately, the ideal treatments of Aims 2 and 3 will be combined to produce a workable therapy that will block soma death and stimulate it to regenerate a functional axon. The findings of Aim 1 will also enhance our treatment of glaucoma by helping to identify the genetic basis of the variable susceptibility to optic nerve damage at the level of Bax gene function.